Acrylamide from Maillard reaction products

Acrylamide formed
(µmol per mol amino acid)
e-mail: d.s.mottram@reading.ac.uk
†Procter Department of Food Science, University of
Leeds, Leeds LS2 9JT, UK
1. Rosen, J. & Hellenas, K.-E. Analyst 127, 880–882 (2002).
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Competing financial interests: declared none.
Food chemistry
Acrylamide from Maillard
reaction products
The discovery of the adventitious formation
of the potential cancer-causing
agent acrylamide in a variety of foods
during cooking 1,2 has raised much concern 3,4 ,
but the chemical mechanism(s) governing
its production are unclear. Here we show
that acrylamide can be released by the
thermal treatment of certain amino acids
(asparagine, for example), particularly in
combination with reducing sugars, and of
early Maillard reaction products (N-glycosides)
5 . Our findings indicate that the
Maillard-driven generation of flavour and
colour in thermally processed foods can —
under particular conditions — be linked to
the formation of acrylamide.
We heated 20 amino acids individually
at 180 °C for 30 min and found that acrylamide
is formed under these conditions
from methionine and from asparagine
(3.651.4 and 0.5650.05 µmol acrylamide
per mol amino acid, respectively; all results
are averages of n46 independent determinations
unless stated otherwise).
When pyrolysed at 180 °C with an
equimolar amount of glucose, asparagine in
particular generates significant amounts of
acrylamide, reaching an average of
368 m mol mol 11 after an incubation time (t i )
of 30 min. If asparagine monohydrate was
used in the incubation or water was added to
the reaction (0.05 ml) before thermolysis, the
release of acrylamide was enhanced nearly
threefold (9605210 m mol mol 11 ), or over
1,700 times the amount formed from
asparagine alone under the same conditions.
Reaction of methionine and glutamine
with equimolar amounts of glucose at
180 °C also increased the formation of
acrylamide, which occurred rapidly in each
case (t i 45 min; Fig. 1a). Cysteine was found
to liberate acrylamide after condensation
with glucose (2.050.8 m mol mol 11 at
t i 430 min and 180 °C).
Investigating the role of different carbohydrates
in the formation of acrylamide, we
found that pyrolysing any of these amino
acids (Asn, Gln, Met, Cys) with an equimolar
amount of D-fructose, D-galactose, lactose or
sucrose all led to a significant release of acrylamide,
with comparable yields from each
sugar. No acrylamide was detected when any
of these carbohydrates was heated alone.
To test whether early Maillard products
such as N-glycosides could be acrylamide
precursors in thermal decomposition reactions,
we measured the yields of acrylamide
after pyrolysis (t i 420 min, 180 °C) of
0.2 mmol of four different N-glycosides
(Fig. 1b). Yields were significant (in m mol
per mol N-glycoside: compound 1,
1,3055323; 2, 1,4195278; 3,1452.7; and 4,
8.151.5) and comparable to those released
from the amino-acid and reducing-sugar
precursors under the same
conditions. Furthermore, compound 1 was
confirmed as an intermediate in the
asparagine/glucose reaction by high-resolution
mass-spectrometric analysis of a
methanol extract of the pyrolysate.
On the basis of structural considerations,
asparagine or the N-glycosides 1 and
2 could be direct precursors of acrylamide
under pyrolytic conditions. Condensation
of asparagine with 13 C 6 -labelled glucose
confirmed that the amino acid is the carbon
source of acrylamide. Upon pyrolysis, formation
of the corresponding N-glycoside
probably facilitates the decarboxylation
step and heterolytic cleavage of the nitrogen–carbon
bond to liberate acrylamide
(CH 2 5CHCONH 2 ). Although decarboxylation
is favoured at higher temperatures, the
N-glycosidic bond seems to facilitate the
deamination step.
Further evidence to support this pathway
to acrylamide production is provided by the
98.6% incorporation of nitrogen-15 label
into acrylamide after the pyrolysis of 15 N-
amide-labelled asparagine with glucose; there
was no incorporation into acrylamide when
15 N-a-amino-labelled asparagine was used in
the same reaction. Results from similar isotope-labelling
experiments (not shown) to
determine the route of acrylamide formation
from different N-glycosides produced by glucose
pyrolysis with glutamine or methionine
are less clear-cut, which suggests that other
pathways (such as that for homolytic cleavage)
might also lead to acrylamide.
The N-glycosidic bond is labile in the
presence of water 6 or under acidic and neutral
pH conditions 7 , hydrolysing rapidly to
the reducing sugar and amino acid. At higher
pH, however, N-glycosides can be isolated as
bimolecular complexes in the presence of
polyvalent alkaline or transition-metal ions 8 .
In food-processing systems that incorporate
conditions of high temperature and water
loss, N-glycoside formation could be
favoured; when this condensation occurs
between reducing sugars and certain amino
acids, a direct pathway is opened up to
brief communications
a
b
HO
HO
HO
HO
10,000
HO
HO
HO
HO
1,000
100
10
1
0.1
5 10 20 30 60
Time (min) at 180 °C
OH
O
1
2
OH
NH NH 2
Figure 1 Production of acrylamide from N-glycosides. a, Logarithmic-scale
plot of the formation of acrylamide over time in
pyrolysates of glucose with glutamine (triangles), asparagine
(squares) or methionine (circles). Each data point represents the
average of n43 independent determinations; the coefficient of
variation was less than 25%. For acrylamide analysis (by liquid
chromatography coupled to electrospray ionization tandem mass
spectrometry), pyrolysates were supplemented with 13 C 3 -acrylamide
(50 ng), then suspended in hot water (more than 90 7C),
sonicated and filtered before being applied to a solid-phase
extraction cartridge (OASIS HLB, 0.2 g). Acrylamide eluted with
20% methanol was separated on a Shodex RSpak DE-613
polymer column with isocratic solvent flow. Detection by mass
spectrometry was in the multiple-reaction monitoring mode with
the characteristic fragmentation transitions for acrylamide (m/z
72➝55, 72➝27, 72➝54) and confirmed by ion ratios (55/54
and 55/27). Further details are available from the authors.
b, Chemical structures of the potassium salts of N-(D-glucos-1-yl)-
L-asparagine (1), N-(D-fructos-2-yl)-L-asparagine (2), N-(D-glucos-
1-yl)-L-glutamine (3) and N-(D-glucos-1-yl)-L-methionine (4).
potential progenitors of acrylamide.
Richard H. Stadler, Imre Blank, Natalia
Varga, Fabien Robert, Jörg Hau, Philippe A.
Guy, Marie-Claude Robert, Sonja Riediker
Nestlé Research Center, Nestec, Vers-chez-les-Blanc,
1000 Lausanne 26, Switzerland
e-mail: richard.stadler@rdls.nestle.com
1. Swedish National Food Agency website http://www.slv.se
2. Rosén, J. & Hellenäs, K.-E. The Analyst 127, 880–882 (2002).
3. WHO FAO/WHO Consultation on the Health Implications of
Acrylamide in Food (Geneva, 25–27 June 2002)
1
4
CO 2 K
O
2 NH NH
OH
2
HO 1 CO 2 K O
2
OH
O
O
1
2
OH
NH
NH 2
3
CO 2 K
OH
O
1
2
OH
NH
S
Me
CO 2 K
O
NATURE | VOL 419 | 3 OCTOBER 2002 | www.nature.com/nature 449
© 2002 Nature Publishing Group

ief communications
http://www.who.int/fsf/
4. European Commission Scientific Committee on Food (SCF)
Opinion of the Scientific Committee on Food on New Findings
Regarding the Presence of Acrylamide in Food
(SCF/CS/CNTM/CONT/4 Final, 3 July 2002)
http://europa.eu.int/comm/food/fs/scf/index_en.html
5. Ledl, F. & Schleicher, E. Angew. Chem. Int. Ed. Engl. 29,
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Chemistry and Biochemistry (eds Pigman, W. & Hortin, D.)
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459–464 (1989).
Competing financial interests: declared none.
Quantum cryptography
A step towards global
key distribution
Large random bit-strings known as
‘keys’ are used to encode and decode
sensitive data, and the secure distribution
of these keys is essential to secure communications
across the globe 1 . Absolutely
secure key exchange 2 between two sites has
now been demonstrated over fibre 3 and
free-space 4–6 optical links. Here we describe
the secure exchange of keys over a freespace
path of 23.4 kilometres between two
mountains. This marks a step towards
accomplishing key exchange with a near-
Earth orbiting satellite and hence a global
key-distribution system.
The security of our key-exchange system
is guaranteed by encoding single photons
using two sets of orthogonal polarizations.
Our transmitter module (Alice; Fig. 1)
incorporates a miniature source of polarization-coded
faint pulses (approximating
single photons; C.K., P.Z., M.H. and H.W.,
unpublished results), where 0° or 45° polarization
encode binary zero, and 90° or 135°
code binary one. These light pulses are
expanded and collimated in a simple
telescope to a beam of about 50 mm and
then accurately aligned on the receiver
(Bob; Fig. 1), a 25-cm-diameter commercial
telescope. Light is collected and focused
onto a compact four-detector photoncounting
module (Fig. 1). A detection in
any one detector then has an associated bit
value, measurement basis (0° or 45°) and
detection time. The bit values then form a
raw key string. Valid bits are measured in
the same basis as that in which they were
encoded.
Alice and Bob use a standard communications
channel, such as a mobile telephone,
to ascertain which bits arrived (many are
lost) and which measurement basis was used,
then they both discard the invalid bits —
which leaves them with nearly identical
random bit-strings, the sifted key. Eavesdropping
measurements on the single
photons disturb the encoding and introduce
errors of up to 25%, so Alice and Bob test for
errors in a short section of sifted key to
Alice
Zugspitze
(2,950 m)
Computer
Fast pulse
generator
Alice
Figure 1 Overview of the experiment against a relief map of the trial site. In the Alice module, four separate lasers (LDs) encode the four
polarizations based on a random bit-string fed from the Alice computer. They are combined in a spatial filter (A,A) using a conical mirror
(M) and a lens (L). The beam expands to 50 mm and is collimated in an output lens (L8). In the Bob module, a telescope (T) collects the
light, which is filtered (F) and then spilt in a polarization-insensitive beam-splitter (BS), passing on to polarizing beam-splitters (PBS) and
four photon-counting detectors (D). One polarizing beam-splitter is preceded by a 45° polarization rotator (R). A click in one of the photoncounting
detectors D(u, B) sets the bit value B and the measurement basis u.
verify the security of the channel. Low error
rates due to background light detection and
polarization settings are securely eliminated
by using classical error-correcting codes sent
over the mobile-telephone link.
In the long-range experiment, Alice was
located at a small experimental facility on
the summit of Zugspitze in southern
Germany, and Bob was on the neighbouring
mountain of Karwendelspitze, 23.4 km
away. At this distance, the transmitted
beam was 1–2 m in diameter and was
only weakly broadened by air-turbulence
effects at this altitude. Lumped optical losses
of about 18–20 decibels were measured
and, using faint pulses containing 0.1
photons per bit, the detected bit rate at
Bob was 1.5–2 kilobits per second (receiver
efficiency of 15%).
Operating at night with filters of 10-nm
bandwidth reduced the background
counts, and errors appeared in less than
5% of key bits. After sifting and error
correction, net key exchange rates were
hundreds of bits per second. In a series of
experiments, several hundreds of kilobits
of identical key string were generated at
Alice and Bob.
In associated experiments in poorer visibility,
we showed that key exchange could
be carried out when transmission losses
were up to 27 decibels, but improvements
in receiver efficiency and background
counts should take us beyond 33 decibels.
With this performance, key exchange to
near-Earth orbit (500–1,000 km range)
should become possible.
Until now, the principal method of
high-security key exchange has been the
M
4 LDs
S
A
A
L
L'
Bob
Westliche
karwendespitze
(2,244 m)
23.4 km
Mobile phone
link
T F BS R
Bob
PBS
D(0°,1)
D(0°,0)
PBS
D(45°,0)
D(45°,1)
Computer
‘trusted courier’ carrying a long random
bit-string, the key, from one location to the
other. Our experiment paves the way for
the development of a secure global keydistribution
network based on optical links
to low-Earth-orbit satellites. We note that a
10-kilometre key-exchange experiment has
recently been announced 7 .
C. Kurtsiefer*, P. Zarda*, M. Halder*,
H. Weinfurter*, P. M. Gorman†,
P. R. Tapster †, J. G. Rarity†
*Ludwig-Maximilian University, 80799 Munich,
Germany
†Photonics Department, QinetiQ, Malvern ,
Worcestershire WR14 3PS, UK
e-mail: jgrarity@qinetiq.com
1. Singh, S. The Code Book (Anchor, New York, 1999).
2. Bennett, C. H. et al. J. Cryptol. 5, 3–28 (1992).
3. Gisin, N., Ribordy, G., Tittel, W. & Zbinden, H. Rev. Mod. Phys.
74, 145–196 (2002).
4. Buttler, W. T. et al. Phys. Rev. Lett. 84, 5652–5655 (2000).
5. Rarity, J. G., Gorman, P. M. & Tapster, P. R. Electron. Lett.
37, 512–514 (2001).
6. Rarity, J. G., Gorman, P. M. & Tapster, P. R. J. Mod. Opt.
48, 1887–1901 (2001).
7. Hughes, R. J., Nordholt, J. E., Derkacs, D. & Peterson, C. G.
New J. Phys. 4, 43.1–43.14 (2002).
Competing financial interests: declared none.
erratum
Cognitive change and the APOE ;4 allele
I. J. Deary, M. C. Whiteman, A. Pattie, J. M. Starr,
C. Hayward, A. F. Wright, A. Carothers, L. J. Whalley
Nature 418, 932 (2002)
In the second sentence of the seventh paragraph of this
communication, the MMSE scores are incorrectly specified
as less than or equal to 28; these should read as
greater than or equal to 28.
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